Structural requirements for high affinity ligand binding by estrogen receptors: a comparative analysis of truncated and full length estrogen receptors expressed in bacteria, yeast, and mammalian cells.

Published

Journal Article

In order to better understand the structural requirements for effective high affinity binding of estrogens and antiestrogens by the human estrogen receptor (ER), a comparative study was undertaken in which we examined: 1) native ER from the MCF-7 ER-positive human breast cancer cell line; 2) full length ER expressed in yeast; 3) the ER hormone binding domain (amino acid residues 302-595) expressed in yeast; 4) a bacterially expressed protein A fusion product encoding a truncated ER (amino acid residues 240-595); and 5) a synthetic peptide encompassing amino acids 510-551 of the ER. The binding parameters studied included affinity, kinetics, structural specificity for ligands, and stability. Full length ER expressed in yeast was very similar to the MCF-7 ER in its affinity [dissociation constant (Kd), 0.35 +/- 0.05 nM], dissociation rate (t1/2, 3-4 h at 25 C), and structural specificity for both reversible and covalently attaching affinity ligands. While the truncated ER expressed in yeast was similar to MCF-7 ER in its specificity of ligand binding, it showed a slightly reduced affinity for estradiol (Kd, 1.00 +/- 0.17 nM). The bacterially expressed ER also had a lower affinity for estradiol (Kd, 1.49 +/- 0.16 nM), which may be due in part to an increase in the dissociation rate (t1/2, 0.5 h at 25 C). The attachment of covalent affinity ligands and structural specificity for a variety of reversible ligands was comparable in the bacterially expressed ER to that observed for the receptors expressed in MCF-7 cells and yeast.(ABSTRACT TRUNCATED AT 250 WORDS)

Full Text

Duke Authors

Cited Authors

  • Wooge, CH; Nilsson, GM; Heierson, A; McDonnell, DP; Katzenellenbogen, BS

Published Date

  • June 1992

Published In

Volume / Issue

  • 6 / 6

Start / End Page

  • 861 - 869

PubMed ID

  • 1495491

Pubmed Central ID

  • 1495491

International Standard Serial Number (ISSN)

  • 0888-8809

Digital Object Identifier (DOI)

  • 10.1210/mend.6.6.1495491

Language

  • eng

Conference Location

  • United States